Project Summary The goal of this project is to understand the role of somatic mutations in neurodegeneration, specifically in neurodegenerative disorders Alzheimer's Disease (AD) and Ataxia Telangiectasia (AT), using single-cell analysis. AD is a progressive neurodegenerative disorder leading to the loss of memory and other cognitive functions; AT is another neurodegenerative disorder with inherited defects in DNA double strand break (DSB) repair. Somatic mutations have been studied most extensively in cancer, but they also cause neurodevelopmental disorders such as epilepsy and hemimegalencephaly. Previous studies have found somatic mosaic mutations in causally implicated genes in some neurodegenerative diseases, such as presenilin-1 in AD. Other studies, including our work, have suggested that the lack of proper DSB repair in AT may allow increased accumulation of somatic mutations, including those in repetitive DNA, and lead to neuronal cell death. Our recent single-neuron genomic studies showed that even neurologically normal human brains are a patchwork of somatic mutations that occur throughout one's life, and that active transcription may play a role in generation of rare somatic mutations. We therefore hypothesize that late-development or even post-mitotic mutations in a small number of neurons may have a functional role in the loss of synaptic function and cell loss in neurodegenerative diseases. To overcome the limited detection sensibility for low frequency variants in current bulk-cell analysis, we will employ single-cell genomics to test our hypothesis, using novel computational tools to circumvent the noise in the data introduced by the genome amplification process necessary in current single-cell sequencing protocols. In Aim 1, we propose to develop robust analytical methods to mitigate the impact of genome amplification bias in detecting multiple forms of somatic mutations, including repeat aberrations (tandem repeats such as telomere and retrotransposons). In Aim 2, we will analyze somatic mutations in post-mortem brains of patients with AT and AD using single-neuron whole genome sequencing.